This study is engaged in the detection and characterization of transient oxygenated intermediated which occur in biological oxygenases, with the hope that this will help us to understand the mechanism of the biological activation of molecular oxygen. We shall continue the studies on the nonheme iron protein, protocatechuate dioxygenase, with the aim of indentifying the transient intermediates which we have previously observed in the oxygen reactions. We shall have two main approaches: (a) A study of the proton flux, in order to establish at which stages of the reaction protons are releases or taken up, and (b) a study of the EPR properties of these intermediates, using rapid freeze quench technique, may help to determine whether any major iron-ligand changes occur in the course of the reaction. A new study of a similar enzyme, catechol oxygenase, will begin this year. It is hoped that parallel studies of similar enzymes will complement each other. We shall also continue the kinetic and spectral studies of liver microsomal amine oxidase. We plan to compare this reaction wth other flavoprotein hydroxylases. Collaborative efforts will also continue with Drs. Vincent Massey, M. J. Coon, D. E. Hultquist, and J. Fee, all of the Department of Biological Chemistry.